CD4 T cells are the key immunoregulatory cells in immunoregulatory responses. They are necessary for development of Ab-secreting B cells and cytolytic effector CD8 cells and they play critical role sin activating macrophages and carrying out inflammatory responses. When an animal is immunized by encountering a pathogen or by vaccination, CD4 memory T cells develop and are critical to the "memory" state of the host, which results in a much faster, greatly more effective response upon re- encounter with an infectious organism, so that it is destroyed before any pathology develops. Thus, an understanding of the factors regulation the generation of CD4 memory and any of their behavior and how best to elicit their response, is critical to understanding immunity and designing vaccines and strategies for vaccination. The objectives of this project are to define the factors which promote the generation and maintenance of memory CD4 T cells and to define the behavior of memory cells. We will employ strains of mice transgenic for T cell receptors, vial dyes and allelic and genetic markers, to visualize the in vivo generation of memory CD4 T cells and their subsequent responses to rechallenge. In Aim 1 we will take naive, antigen-inexperienced, CD4 T cells, and stimulate them in vitro to see which factors and manipulations render them competent to become memory cells after transfer to adoptive hosts in the absence of antigen. We will test the roles of antigen dose, co-stimulatory factors and growth-survival cytokines on the transition of naive CD4 T cells to pre- memory cells. In Aim 2, we will study the fate of transferred pre-memory CD4 T cells to identify steps in the transition to memory and continue our studies of the properties associated with that transition. To define the molecules and mechanisms likely to be required for this transition in Aim 3, we will vary the hosts, using mice deficient in particular the relevant molecules or we will introduce blocking Ab to such molecules into hosts. We will focus on such host factors as cytokines, chemokines and cell:cell interaction molecules which may provide the signals for the transition. In this Aim we will collaborate extensively with Project 3 which will study the role of the many of the same molecules in T cell homing, recirculation and recruitment into inflammatory sites. Finally in Aim 4 we will visualize the response of memory cells in situ and examine what factors regulate the memory CD4 response, focusing on possible differences between requirements for elicitation of naive and memory CD4 T cells. Throughout we will directly compare CD4 and CD8 T cells in collaboration with Project 2, to reach an understanding of how these functionally diverse subsets are differentially regulated. These studies with the studies in Project on CD8 memory and Project 3 on the regulation of both CD4 and CD8 T cell migration will provide important insights into the mechanisms and factors which regulate the generation of T cell memory and the behaviors of memory T cells which determine immunity.